Publications Resulting from ISS National Lab Sponsored Projects

The attenuation of sedimentation and convection in microgravity can sometimes decrease irregularities formed during macromolecular crystal growth. Current terrestrial protein crystal growth (PCG) capabilities are very different than those used during the Shuttle era and that are currently on the International Space Station (ISS). The focus of this experiment was to demonstrate the use of a commercial off-the-shelf, high throughput, PCG method in microgravity. Using Protein BioSolutions’ microfluidic Plug Maker™/CrystalCard™ system, we tested the ability to grow crystals of the regulator of glucose metabolism and adipogenesis: peroxisome proliferator-activated receptor gamma (apo-hPPAR-γ LBD), as well as several PCG standards. Overall, we sent 25 CrystalCards™ to the ISS, containing ~10,000 individual microgravity PCG experiments in a 3U NanoRacks NanoLab (1U = 103 cm.). After 70 days on the ISS, our samples were returned with 16 of 25 (64%) microgravity cards having crystals, compared to 12 of 25 (48%) of the ground controls. Encouragingly, there were more apo-hPPAR-γ LBD crystals in the microgravity PCG cards than the 1g controls. These positive results hope to introduce the use of the PCG standard of low sample volume and large experimental density to the microgravity environment and provide new opportunities for macromolecular samples that may crystallize poorly in standard laboratories.

?Musica universalis? is an ancient philosophical concept claiming the movements of celestial bodies follow mathematical equations and resonate to produce an inaudible harmony of music, and the harmonious sounds that humans make were an approximation of this larger harmony of the universe. Besides music, electromagnetic waves such as light and electric signals also are presented as harmonic resonances. Despite the seemingly universal theme of harmonic resonance in various disciplines, it was not until recently that the same harmonic resonance was discovered also to exist in biological systems. Contrary to traditional belief that a biological system is either at stead-state or cycles with a single frequency, it is now appreciated that most biological systems have no homeostatic ?set point,? but rather oscillate as composite rhythms consisting of superimposed oscillations. These oscillations often cycle at different harmonics of the circadian rhythm, and among these, the ~12-hour oscillation is most prevalent. In this review, we focus on these 12-hour oscillations, with special attention to their evolutionary origin, regulation, and functions in mammals, as well as their relationship to the circadian rhythm. We further discuss the potential roles of the 12-hour clock in regulating hepatic steatosis, aging, and the possibility of 12-hour clock?based chronotherapy. Finally, we posit that biological rhythms are also musica universalis: whereas the circadian rhythm is synchronized to the 24-hour light/dark cycle coinciding with the Earth?s rotation, the mammalian 12-hour clock may have evolved from the circatidal clock, which is entrained by the 12-hour tidal cues orchestrated by the moon.

β-blockers are unsuccessful in eliminating stress-induced ventricular arrhythmias in approximately 25% of patients with catecholaminergic polymorphic ventricular tachycardia (CPVT). Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) generated from these patients have potential for investigating the phenomenon, but it remains unknown whether they can recapitulate patient-specific drug responses to β-blockers. This study assessed whether the inadequacy of β-blocker therapy in an individual can be observed in vitro using patient-derived CPVT iPSC-CMs. A CPVT patient harboring a novel mutation in the type 2 cardiac ryanodine receptor (RyR2) was identified whose persistent ventricular arrhythmias during β-blockade with nadolol were abolished during flecainide treatment. iPSC-CMs generated from this patient and two control individuals expressed comparable levels of excitation-contraction genes, but assessment of the sarcoplasmic reticulum Ca2+ leak and load relationship revealed intracellular Ca2+ homeostasis was altered in CPVT iPSC-CMs. β-adrenergic stimulation potentiated spontaneous Ca2+ waves and unduly frequent, large, and prolonged Ca2+ sparks in CPVT compared to control iPSC-CMs, validating the disease phenotype. Pursuant to the patient's in vivo responses, nadolol treatment during β-adrenergic stimulation achieved negligible reduction of Ca2+ wave frequency and failed to rescue Ca2+ spark defects in CPVT iPSC-CMs. In contrast, flecainide reduced both frequency and amplitude of Ca2+ waves and restored the frequency, width, and duration of Ca2+ sparks to baseline levels. By recapitulating a CPVT patient's improved response to flecainide compared to β-blocker therapy in vitro, these data provide new evidence that iPSC-CMs can capture basic components of patient-specific drug responses.

This paper describes a novel miniature microcontroller based curve tracing circuit, which was designed to monitor the environmental effects on Silicon Carbide Junction Field Effect Transistor (SiC JFET) device performance, while exposed to the low earth orbit environment onboard the International Space Station (ISS) as a resident experiment on the 7th Materials on the International Space Station Experiment (MISSE7). Specifically, the microcontroller circuit was designed to operate autonomously and was flown on the external structure of the ISS for over a year. This curve tracing circuit is capable of measuring current vs. voltage (I-V) characteristics of transistors and diodes. The circuit is current limited for low current devices and is specifically designed to test high temperature, high drain-to-source resistance SiC JFETs. The results of each I-V data set are transmitted serially to an external telemetered communication interface. This paper discusses the circuit architecture, its design, and presents example results.

Bacteria behave differently in space, as indicated by reports of reduced lag phase, higher final cell counts, enhanced biofilm formation, increased virulence, and reduced susceptibility to antibiotics. These phenomena are theorized, at least in part, to result from reduced mass transport in the local extracellular environment, where movement of molecules consumed and excreted by the cell is limited to diffusion in the absence of gravity-dependent convection. However, to date neither empirical nor computational approaches have been able to provide sufficient evidence to confirm this explanation. Molecular genetic analysis findings, conducted as part of a recent spaceflight investigation, support the proposed model. This investigation indicated an overexpression of genes associated with starvation, the search for alternative energy sources, increased metabolism, enhanced acetate production, and other systematic responses to acidity all of which can be associated with reduced extracellular mass transport.

Understanding and quantifying the natural processes that occur along coasts are critical components of managing environmental resources and planning and executing coastal operations, from humanitarian relief to military actions. However, the coastal ocean is complicated, with dissolved and suspended matter that hinders water transparency, phytoplankton blooms that can be toxic, and bathymetry and bottom types that vary over spatial scales of tens of meters, all of which affect processes in an area that spans millions of square kilometers. A hyperspectral imager collects the spectrum of the light received from each pixel in an image. For environmental characterization the wavelength range typically spans the visible and shortwave infrared wavelengths, and the spectrum is collected in contiguous spectral intervals 1–10 nanometers wide. This spectral information is exploited to provide significantly more information about vegetation, minerals, and other components in the scene than can be retrieved from panchromatic or even multispectral imagery, which rely primarily on the shape of the object for detection [Goetz et al., 1985]. Such technology can also work over shallow seas. Over the past 2 decades, experiments with hyperspectral imagers on airborne platforms have demonstrated the ability to characterize the coastal environment [Davis et al., 2002, Davis et al. 2006] and produce maps of coastal bathymetry, in‐water constituents, and bottom type.

An ongoing challenge in biomedical research is the search for simple, yet robust assays using 3D cell cultures for toxicity screening. This study addresses that challenge with a novel spheroid assay, wherein spheroids, formed by magnetic 3D bioprinting, contract immediately as cells rearrange and compact the spheroid in relation to viability and cytoskeletal organization. Thus, spheroid size can be used as a simple metric for toxicity. The goal of this study was to validate spheroid contraction as a cytotoxic endpoint using 3T3 fibroblasts in response to 5 toxic compounds (all-trans retinoic acid, dexamethasone, doxorubicin, 5′-fluorouracil, forskolin), sodium dodecyl sulfate (+control), and penicillin-G (−control). Real-time imaging was performed with a mobile device to increase throughput and efficiency. All compounds but penicillin-G significantly slowed contraction in a dose-dependent manner (Z’ = 0.88). Cells in 3D were more resistant to toxicity than cells in 2D, whose toxicity was measured by the MTT assay. Fluorescent staining and gene expression profiling of spheroids confirmed these findings. The results of this study validate spheroid contraction within this assay as an easy, biologically relevant endpoint for high-throughput compound screening in representative 3D environments.

The Hyperspectral Imager for the Coastal Ocean (HICO) instrument currently on board the International Space Station is a new sensor designed specifically for the studies of turbid coastal waters and large inland lakes and rivers. It covers the wavelength range between 0.4 and 0.9 μm with a spectral resolution of 5.7 nm and a spatial resolution of approximately 90 m. The HICO sensor is not equipped with a second-order blocking filter in front of the focal plane array. As a result, the second-order light from the shorter visible spectral region falls onto the detectors covering the near-IR spectral region above 0.8 μm. In order to have accurate radiometric calibration of the near-IR channels, the second-order light contribution needs to be removed. The water-leaving radiances of these near-IR channels over clear ocean waters are close to zero because of strong liquid water absorption above 0.8 μm. Through analysis of HICO imaging data containing features of shallow underwater objects, such as coral reefs, we have developed an empirical technique to correct for the second-order light effects in near-IR channels. HICO data acquired over Midway Island in the Pacific Ocean and the Bahamas Banks in the Atlantic Ocean are used to demonstrate the effectiveness of the new technique.

Microgravity facilitates the opportunistic infections by augmenting the pathogenic virulence and suppressing the host resistance. Hence the extraterrestrial infections may activate potentially novel bionetworks different from the terrestrial equivalent, which could only be probed by investigating the host-pathogen relationship with a minimum of terrestrial bias.

Huntington's disease is caused by expansion of a polyglutamine (polyQ) repeat in the huntingtin protein. A structural basis for the apparent transition between normal and disease-causing expanded polyQ repeats of huntingtin is unknown. The "linear lattice" model proposed random-coil structures for both normal and expanded polyQ in the preaggregation state. Consistent with this model, the affinity and stoichiometry of the anti-polyQ antibody MW1 increased with the number of glutamines. An opposing "structural toxic threshold" model proposed a conformational change above the pathogenic polyQ threshold resulting in a specific toxic conformation for expanded polyQ. Support for this model was provided by the anti-polyQ antibody 3B5H10, which was reported to specifically recognize a distinct pathologic conformation of soluble expanded polyQ. To distinguish between these models, we directly compared binding of MW1 and 3B5H10 to normal and expanded polyQ repeats within huntingtin exon 1 fusion proteins. We found similar binding characteristics for both antibodies. First, both antibodies bound to normal, as well as expanded, polyQ in huntingtin exon 1 fusion proteins. Second, an expanded polyQ tract contained multiple epitopes for fragments antigen-binding (Fabs) of both antibodies, demonstrating that 3B5H10 does not recognize a single epitope specific to expanded polyQ. Finally, small-angle X-ray scattering and dynamic light scattering revealed similar binding modes for MW1 and 3B5H10 Fab-huntingtin exon 1 complexes. Together, these results support the linear lattice model for polyQ binding proteins, suggesting that the hypothesized pathologic conformation of soluble expanded polyQ is not a valid target for drug design.

A precision measurement by AMS of the antiproton flux and the antiproton-to-proton flux ratio in primary cosmic rays in the absolute rigidity range from 1 to 450 GV is presented based on 3.49×105 antiproton events and 2.42×109 proton events. The fluxes and flux ratios of charged elementary particles in cosmic rays are also presented. In the absolute rigidity range ∼60 to ∼500  GV, the antiproton ¯p, proton p, and positron e+ fluxes are found to have nearly identical rigidity dependence and the electron e− flux exhibits a different rigidity dependence. Below 60 GV, the (¯p/p), (¯p/e+), and (p/e+) flux ratios each reaches a maximum. From ∼60 to ∼500  GV, the (¯p/p), (¯p/e+), and (p/e+) flux ratios show no rigidity dependence. These are new observations of the properties of elementary particles in the cosmos.

As a demonstrator for technologies for the next generation of ocean color sensors, the Hyperspectral Imager for the Coastal Ocean (HICO) provides enhanced spatial and spectral resolution that is required to understand optically complex aquatic environments. In this study we apply HICO, along with satellite remote sensing and in situ observations, to studies of phytoplankton ecology in a dynamic coastal upwelling environment—Monterey Bay, CA, USA. From a spring 2011 study, we examine HICO-detected spatial patterns in phytoplankton optical properties along an environmental gradient defined by upwelling flow patterns and along a temporal gradient of upwelling intensification. From a fall 2011 study, we use HICO’s enhanced spatial and spectral resolution to distinguish a small-scale “red tide” bloom, and we examine bloom expansion and its supporting processes using other remote sensing and in situ data. From a spectacular HICO image of the Monterey Bay region acquired during fall of 2012, we present a suite of algorithm results for characterization of phytoplankton, and we examine the strengths, limitations, and distinctions of each algorithm in the context of the enhanced spatial and spectral resolution.

There is a significant need for in vitro methods to study drug-induced liver injury that are rapid, reproducible, and scalable for existing high-throughput systems. However, traditional monolayer and suspension cultures of hepatocytes are difficult to handle and risk the loss of phenotype. Generally, three-dimensional (3D) cell culture platforms help recapitulate native liver tissue phenotype, but suffer from technical limitations for high-throughput screening, including scalability, speed, and handling. Here, we developed a novel assay for cytochrome P450 (CYP450) induction/inhibition using magnetic 3D cell culture that overcomes the limitations of other platforms by aggregating magnetized cells with magnetic forces. With this platform, spheroids can be rapidly assembled and easily handled, while replicating native liver function. We assembled spheroids of primary human hepatocytes in a 384-well format and maintained this culture over five days, including a 72 h induction period with known CYP450 inducers/inhibitors. CYP450 activity and viability in the spheroids were assessed and compared in parallel with monolayers. CYP450 activity was induced/inhibited in spheroids as expected, separate from any toxic response. Spheroids showed a significantly higher baseline level of CYP450 activity and induction over monolayers. Positive staining in spheroids for albumin and multidrug resistance-associated protein (MRP2) indicates the preservation of hepatocyte function within spheroids. The study presents a proof-of-concept for the use of magnetic 3D cell culture for the assembly and handling of novel hepatic tissue models.

Several ocean color earth observation satellite sensors are presently collecting daily imagery, including the Hyperspectral Imager for the Coastal Ocean (HICO). HICO has been operating aboard the International Space Station since its installation on September 24, 2009. It provides high spatial resolution hyperspectral imagery optimized for the coastal ocean. Atmospheric correction, however, still remains a challenge for this sensor, particularly in optically complex coastal waters. In this paper, we assess the application of the cloud-shadow atmospheric correction approach on HICO data and validate the results with the in situ data. We also use multiple sets of cloud, shadow, and sunlit pixels to correct a single image multiple times and intercompare the results to assess variability in the retrieved reflectance spectra. Retrieved chlorophyll values from this intercomparison are similar and also agree well with the in situ chlorophyll measurements.

This letter focuses on water-quality estimation in the northern Adriatic Sea using physically-based methods applied to image obtained with the Hyperspectral Imager for the Coastal Ocean (HICO™). Optical properties of atmosphere and water were synchronously measured to parameterise such methods. HICO™-derived maps of chlorophyll-a (chl-a) and suspended particulate matter (SPM) indicated low values, in the range of 0-3 mg m-3 and 0-4 g m-3, respectively, correlating significantly with field data (R2 = 0.71 for chl-a and R2 = 0.85 for SPM). The results, on analysis, identify clear waters in the open sea and moderately turbid waters near the coast due to river sediment discharge and organic matter from coastal lagoons. These findings support the use of HICO™ data to assess water-quality parameters in coastal zones and suggest the feasibility of integrating them with future-generation space-borne hyperspectral images.

Interest in space habitation has grown dramatically with planning underway for the first human transit to Mars. Despite a robust history of domestic and international spaceflight research, understanding behavioral adaptation to the space environment for extended durations is scant. Here we report the first detailed behavioral analysis of mice flown in the NASA Rodent Habitat on the International Space Station (ISS). Following 4-day transit from Earth to ISS, video images were acquired on orbit from 16- and 32-week-old female mice. Spaceflown mice engaged in a full range of species-typical behaviors. Physical activity was greater in younger flight mice as compared to identically-housed ground controls, and followed the circadian cycle. Within 9–11 days after launch, younger (but not older), mice began to exhibit distinctive circling or ‘race-tracking’ behavior that evolved into a coordinated group activity. Organized group circling behavior unique to spaceflight may represent stereotyped motor behavior, rewarding effects of physical exercise, or vestibular sensation produced via self-motion. Affording mice the opportunity to grab and run in the RH resembles physical activities that the crew participate in routinely. Our approach yields a useful analog for better understanding human responses to spaceflight, providing the opportunity to assess how physical movement influences responses to microgravity.

There is evidence that space flight condition-induced biological damage is associated with increased oxidative stress and extracellular matrix (ECM) remodeling. To explore possible mechanisms, changes in gene expression profiles implicated in oxidative stress and in ECM remodeling in mouse skin were examined after space flight. The metabolic effects of space flight in skin tissues were also characterized. Space Shuttle Atlantis (STS-135) was launched at the Kennedy Space Center on a 13-day mission. Female C57BL/6 mice were flown in the STS-135 using animal enclosure modules (AEMs). Within 3?5 h after landing, the mice were euthanized and skin samples were harvested for gene array analysis and metabolic biochemical assays. Many genes responsible for regulating production and metabolism of reactive oxygen species (ROS) were significantly (p < 0.05) altered in the flight group, with fold changes >1.5 compared to AEM control. For ECM profile, several genes encoding matrix and metalloproteinases involved in ECM remodeling were significantly up-/down-regulated following space flight. To characterize the metabolic effects of space flight, global biochemical profiles were evaluated. Of 332 named biochemicals, 19 differed significantly (p < 0.05) between space flight skin samples and AEM ground controls, with 12 up-regulated and 7 down-regulated including altered amino acid, carbohydrate metabolism, cell signaling, and transmethylation pathways. Collectively, the data demonstrated that space flight condition leads to a shift in biological and metabolic homeostasis as the consequence of increased regulation in cellular antioxidants, ROS production, and tissue remodeling. This indicates that astronauts may be at increased risk for pathophysiologic damage or carcinogenesis in cutaneous tissue.

Although bone has remarkable regenerative capacity, about 10% of long bone fractures and 25% to 40% of vertebral fusion procedures fail to heal. In such instances, a scaffold is employed to bridge the lesion and accommodate osteoprogenitors. Although synthetic bone scaffolds mimic some of the characteristics of bone matrix, their effectiveness can vary because of biological incompatibility. Herein, we demonstrate that a composite prepared with osteogenically enhanced mesenchymal stem cells (OEhMSCs) and their extracellular matrix (ECM) has an unprecedented capacity for the repair of critical‐sized defects of murine femora.Furthermore, OEhMSCs do not cause lymphocyte activation, and ECM/OEhMSC composites retain their in vivo efficacy after cryopreservation. Finally, we show that attachment to the ECM by OEhMSCs stimulates the production of osteogenic and angiogenic factors. These data demonstrate that composites of OEhMSCs and their ECM could be utilized in the place of autologous bone graft for complex orthopedic reconstructions.

Two experiments were conducted aboard the International Space Station (ISS) in 2008 and 2009 that engaged elementary and middle school teachers and students worldwide in authentic science investigations designed to increase student knowledge of, and interest in, biology and space life science studies and biomedical careers. In the first project, a pilot called Butterflies and Spiders in Space, 1,876 middle school students tested a protocol for comparing, at near-real time, the behaviors of orb-weaver spiders and painted lady butterflies living in microgravity (aboard ISS) to those of comparable subjects in students' classrooms. Teachers reported that, as a result of project activities, 33% of their students designed additional experiments and 80% of students expressed interest in science careers. The second program, Butterflies in Space, enabled students to observe and investigate the life cycle and behaviors of painted lady butterflies living on ISS, and compare them to butterflies being studied in their own classes. Combining this near real-time experiment with hands-on explorations and webbased instructional strategies, Butterflies in Space reached more than 3,000 teachers, representing an estimated 180,000 students (grades 3-6) or more worldwide. It also received international coverage from a variety of media. Investigators at BioServe Space Technologies of the University of Colorado designed and built the chambers in which the spiders and butterflies were housed on ISS, and led technical and logistical operations for both programs. Baylor College of Medicine (BCM) educators and scientists developed the education framework and managed the web-based distribution of project data and teaching resources.

The nematode Caenorhabditis elegans, a popular organism for biological studies, is being developed as a model system for space biology. The chemically defined liquid medium, C. elegans Maintenance Medium (CeMM), allows axenic cultivation and automation of experiments that are critical for spaceflight research. To validate CeMM for use during spaceflight, we grew animals using CeMM and standard laboratory conditions onboard STS-107, space shuttle Columbia. Tragically, the Columbia was destroyed while reentering the Earth's atmosphere. During the massive recovery effort, hardware that contained our experiment was found. Live animals were observed in four of the five recovered canisters, which had survived on both types of media. These data demonstrate that CeMM is capable of supporting C. elegans during spaceflight. They also demonstrate that animals can survive a relatively unprotected reentry into the Earth's atmosphere, which has implications with regard to the packaging of living material during space flight, planetary protection, and the interplanetary transfer of life.